Promokine pcr mycoplasma test kit 1 c

A HPV-negative, highly aggressive metastatic human tongue squamous cell carcinoma cell line (HSC-3, Japan Health Sciences Foundation, Japan) was cultured with Dulbecco’s Modified Eagle’s Medium (DMEM)/F-12 (Gibco Paisley, UK) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco), 100 µg/ml streptomycin (Gibco), 100 U/ml penicillin (Gibco), 5 µg/ml amphotericin b (Sigma-Aldrich, St. Louis, Missouri, USA), and 50 µg/ml ascorbic acid (PanReac AppliChem, Darmstadt, Germany) in 75 m² flasks. Cells were divided after reaching 80% confluence with 10% trypsin/EDTA (Gibco). The HSC-3 cells were tested negative for mycoplasma using a PromoKine PCR Mycoplasma Test Kit I/C (PromoCell, Heidelberg, Germany). The cell line was authenticated by FIMM Technology Center (Helsinki, Finland).

DNA isolated from human‐derived cell lines with reported wild‐type or mutation status in the oncogenes KRAS and BRAF was used to establish Droplet Digital PCR (ddPCR) assays (Table S1). All cell lines were cultured in media supplemented with 10% heat‐inactivated fetal bovine serum (Thermo Fisher Scientific, Waltham, USA) and 1% penicillin/streptomycin (Biochrom GmbH, Berlin, Germany) at 37°C and 5% CO₂. Prior to DNA isolation, cell lines were tested negative for mycoplasma using the Promokine PCR Mycoplasma Test KIT I/C following manufacturer's specifications (PromoCell GmbH, Heidelberg, Germany). Cell line authenticity was validated by single nucleotide polymorphism profiling with Multiplexion GmbH (Friedrichshafen, Germany). Following manufacturer's instructions, the GeneJET Genomic DNA Purification kit (Thermo Scientific, Waltham, USA) was used to isolate DNA eluted in 100 µL double distilled water.